PROJECT2: MAPPINGTHEPHARMACOGENETICLANDSCAPEFOR PRECISIONMEDICINE SUMMARY Withtherecentoutpouringofsomaticmutationdata,wenowhaveanextensivelistofthegenesinvolvedin cancer. To realize the full potential of this information, we must now use technologies to functionally characterize the mechanisms of how these mutations are integrated to regulate and deregulate cellular networks.Thepremiseofthisprojectisthatsomaticmutationsconvergeintogeneticinteractionnetworks,and these networks bring together mutations of all varieties, including genes with low frequency of oncogenic mutations,andtumorsuppressorprofiles.Wehypothesizethatbysystematicallymappingthesenetworksand quantifyinghowspecificmutationsregulatethesenetworks,newmoleculartargetsforcancertherapycanbe identified. The focus of this proposal is on invasive breast cancer (BC) and head and neck squamous cell carcinomas(HNSCC),diseasesthattogetherresultinover700,000deathseachyearworldwide.Tothisend, the aims of ?Project 2 focus on using stateoftheart highthroughput epistasis mapping and computational approachestosystematicallyinterrogatethefunctionsofindividualgenesandgenepairsinbreastandhead andneckcancerpathology.Coupledwithfunctionalvalidationsinpreclinicalmodelsandclinicaltrialdata,our approachwillresultinunprecedentedinsightsintotheunderlyingtumorbiologyaswellasunravelingofthe geneticvulnerabilitiesofdirecttherapeuticrelevance.Torealizethisgoal,theCCMIwillleveragetechnologies and synergize expertise in the Krogan, Mali, Ideker, Grandis, Gutkind, Ashworth, Mesirov, van ?t Veer and Esserman laboratories. The cornerstone of our approach will be utilization of CRISPRCas9 based reverse geneticscreeningmethodologies,whichourteamhasestablished,thatenablesdenovodiscoveryofgenetic interactions by targeting single or pairs of genes in a highthroughput fashion. Leveraging TCGA data, we proposetointerrogatehighvaluepanelsofthemostfrequentlymutated,amplified,ordeletedgenesinBCand then separately do the same for HNSCC. High throughput screening of the order of10,000interactionsper experimentwillallowustoexhaustivelymaptheunderlyinggeneticinteractionsbetweenthesegenesets(?Aim 1?).Synthesizingthisgeneticinteractiondatawithotherrepositoriesofgeneticinformation,wewillnextbuildan integrative framework to identify highly conserved synthetic lethal interactions (?Aim 2?). We will validate this resource by testing drug sensitivity predictions ?in vitro as well as in existing pharmacogenomic data sets. Finally,tofacilitateclinicaltranslationofthehighestconfidencegenegeneandgenedruginteractions,wewill utilize our extensive library of BC and HNSCC patientderived xenograft (PDX) models to test these interactions?invivo?andwillvalidatetheidentifiedinteractionnetworksinpatienttumormaterialfromtheBC ISPY 2 trial (?Aim 3?). Taken together our highly integrated approach proposes to establish a network of highconfidence,experimentallyvalidated,geneticinteractionstoserveasaresourcetoadvancethepractice ofprecisiononcology.